Jungwoo Lee, Co-Chair, Chemical Engineering
Neil Forbes, Co-Chair, Chemical Engineering
The mucosal barrier in the intestine is vital to maintain selective absorption of nutrients while
protecting internal tissues and maintaining symbiotic relationship with luminal microbiota.
This bio-barrier consists of a cellular epithelial barrier and an acellular mucus
barrier. Secreted mucus regulates barrier function via in situ biochemical and
biophysical interaction with luminal content that continually evolves during digestion
and absorption. Increasing evidence suggests that a mucus barrier is indispensable to maintain
dynamic homeostasis of the gastrointestinal tract. However, the importance of mucus barrier
has been largely underrated for in vitro mucosal tissue modeling. The major gap is
the lack of experimental material (i.e. functional mucins) and platforms to integrate a
relevant thickness of mucus layer with an epithelium under physiological conditions.
Here we report our progress on developing human-relevant micro-physiological models of the mucosal
barrier in static and dynamic settings by using natural mucins derived from a porcine small
intestine (PSI). To overcome limited availability of functional mucus, we first developed a simple
and scalable protocol for natural mucus extraction by directly solubilizing a relatively sterile
inner mucus layer from PSI that is readily accessible. Subsequently, functional separation of mucin
proteins was performed by exploiting pH-dependent reversible sol- gel transition. Under optimized
alkaline condition (0.01M NaOH), the mucus layer was selectively solubilized from the
mucosal surface with a 72% yield (1275 mg/m PSI). The extracted and purified natural mucins
retained essential biophysical and biochemical characteristics. The in vitro mucus barrier
model enabled us to discover ionic (Ca2+) environment dependent mucus barrier and its
The mucus barrier was successfully integrated with human epithelial cell layer (HT-29), which
allowed the studies of bi-directional crosstalk between luminal content and tissue immune cells
through a physiologically relevant mucosal interface. The applied mucus barrier did not
cause any cytotoxic or immunogenic effects to human intestinal and immune cells. As expected,
mucus prevents the transmigration of probiotic bacteria VSL#3. In the absence of mucus, these
bacteria caused epithelial damage, immune cell differentiation and induced production of
pro-inflammatory cytokines IL-8 and TNF-α. The most intriguing result from these studies was that
mucus increased the transmigration of pathogenic Salmonella. Similar to the transmigration of
probiotic bacteria, breach of the mucosal barrier by Salmonella induced production of IL-8 and
TNF-α. The importance of bacterial motility was confirmed by showing that Salmonella with a
knockout that prevents flagella formation does not penetrate the barrier. Co-cultures of
VSL#3 and Salmonella in the mucosal barrier platform demonstrated the differences in
epithelial and immune cell responses under symbiosis or dysbiosis like conditions.
Taking bioengineering approaches, we have developed mucosal barrier models of intestines.
Established models represent cellular and extracellular complexities in a controlled and accessible
manner. We envision that in vitro mucosal barrier models will serve as an enabling tool
for understanding basic biology and disease
progression in the intestines.